The proposal will determine the importance of PFR for peptide and protein immunogenicity. There is increasing interest in developing peptide-based vaccines in order to reduce the possibility of either inducing autoimmunity against a host protein homologous to the immunogen, or inducing an inappropriate immune response that may exacerbate rather than alleviate a disease. The long-term objective of this program is to gain a greater understanding of the influence that peptide flanking residues (PFR) surrounding MHC class II binding epitopes have on T cell responses in general. The first aim of this proposal will be to determine how frequently PFR affect T cell responses and overall immunogenicity. These studies will primarily involve analysis of murine T cell hybridomas and in vivo immunization experiments. The second aim will be to determine the mechanisms controlling how PFR potentiate and/or modulate T cell function. These studies will involve precursor frequency determination, cytokine analysis and proliferation studies to focus on kinetics and magnitude of T cell responses. The methods described herein include those that are common to the laboratory and those that will be new training experiences. This application will establish the immunological significance of PFR and has significant implications for immunoregulation, vaccine development and peptide-based immunotherapy.